Evidence for LINC1-SUN associations at the plant nuclear periphery

Sad1/UNC84 (SUN) domain proteins are a highly conserved family of inner nuclear membrane localised proteins in eukaryotes. One of their main functions is as key components of nucleo-cytoskeletal bridging complexes, in which SUN proteins associate with nucleoskeletal elements. In metazoans these are the lamins, which form a supportive structural network termed the lamina. Plants lack sequence homologs of lamins but have a similar nucleoplasmic structural network to support the plant NE. Putative components of this plant lamina-like structure are Little Nuclei (LINC) proteins, which bear structural resemblance to lamins and fulfil similar functions. This work explores the associations between AtLINC1, AtSUN1 and AtSUN2. AtLINC1 is recruited to the NE by SUN proteins and is immobilised therein. This recruitment and the immobile properties are likely due to AtSUN1/2-AtLINC1 protein interactions occurring in planta. In addition, the SUN N-terminus appears to play an important role in mediating these interactions. The associations between AtLINC1 and plant SUN proteins are a first indicator of how the nucleoskeleton may be anchored to the nuclear membrane in plants. Building on the previous characterisation of Klarsicht/Anc1/Syne1 homology (KASH) like proteins in plants, this study advances the identification and characterisation of nucleo-cytoskeletal bridging complexes in plants

Nuclear envelope dynamics during plant cell division suggest common mechanisms between kingdoms

Behaviour of the NE (nuclear envelope) during open mitosis has been explored extensively inmetazoans, but lack of native markers has limited similar investigations in plants. In the present study, carried out using living synchronized tobacco BY-2 suspension cultures, the non-functional NE marker LBR (lamin B receptor)–GFP (green fluorescent protein) and two native, functional NE proteins, AtSUN1 [Arapidopsis thaliana SUN (Sad1/UNC84) 1] and AtSUN2, we provide evidence that the ER (endoplasmic reticulum)-retention theory for NE membranes is applicable in plants. We also observe two apparently unique plant features: location of the NE-membrane components in close proximity tochromatin throughout division, and spatially distinct reformation of the NE commencing at the chromatin surface facing the spindle poles and concluding at the surface facing the cell plate. Mobility of the proteins was investigated in the interphase NE, during NE breakdown and reformation, in the spindle membranes and the cell plate. A role for AtSUN2 in nuclear envelope breakdown is suggested

The effect of nose geometry on the aerothermodynamic environment of shuttle entry configurations

The effect was studied of nose geometry on the transition criteria for the windward boundary layer, on the extent of separation, on the heat transfer perturbation due to the canopy, and on the surface pressure and the heat transfer in the separated region. The data for each of these problems is analyzed. A literature review that concentrates on separation and the leeward flow-field is presented

Editorial for the SEB Brighton Special Issue: Dynamic organization of the nucleus

Editorial Materia

Identification and characterization of genes encoding the nuclear envelope LINC complex in the monocot species Zea mays

The LINC (Linker of Nucleoskeleton to Cytoskeleton) complex is an essential multi protein structure spanning the nuclear envelope. It connects the cytoplasm to the nucleoplasm, functions to maintain nuclear shape and architecture, and regulates chromosome dynamics during cell division. Knowledge of LINC complex composition and function in the plant kingdom is primarily limited to Arabidopsis, but critically missing from the evolutionarily distant monocots which include grasses, the most important agronomic crops worldwide. To fill this knowledge gap, we identified and characterized 22 maize genes, including a new grass-specific KASH gene family. Using bioinformatic, biochemical, and cell biological approaches, we provide evidence that representative KASH candidates localize to the nuclear periphery and interact with ZmSUN2 in vivo. FRAP experiments using domain-deletion constructs verified that this SUN-KASH interaction was dependent on the SUN but not the coiled-coil domain of ZmSUN2. A summary working model is proposed for the entire maize LINC complex encoded by conserved and divergent gene families. These findings expand our knowledge of the plant nuclear envelope in a model grass species, with implications for both basic and applied cellular research

Absence of SUN1 and SUN2 proteins in Arabidopsis thaliana leads to a delay in meiotic progression and defects in synapsis and recombination

The movement of chromosomes during meiosis involves the location of their telomeres at the inner surface of the nuclear envelope (NE). Sad1/UNC-84 (SUN)-domain proteins are inner NE proteins that are part of complexes linking cytoskeletal elements with the nucleo skeleton, connecting telomeres to the force-generating mechanism in the cytoplasm. These proteins play a conserved role in chromosome dynamics in eukaryotes. Homologues of SUN-domain proteins have been identified in several plant species. In Arabidopsis thaliana two proteins which interact with each other, named AtSUN1 and AtSUN2, have been identified. Immuno localisation with antibodies to AtSUN1 and AtSUN2 proteins revealed that they were associated with the nuclear envelope during meiotic prophase I. Analysis of the double mutant Atsun1-1 Atsun2-2 has revealed severe meiotic defects, namely, a delay in the progression of meiosis, an absence of full synapsis, unresolved interlock-like structures and a reduction in the mean cell chiasma frequency. We propose that in Arabidopsis thaliana, overlapping functions of SUN1 and SUN2 ensure normal meiotic recombination and synapsis

Plants flash their KASH

An Arabidopsis root tip expresses GFP-tagged SINE1 (green), one of many new outer nuclear membrane KASH proteins that Zhou et al. identify in plants. The plant cell walls are labeled with propidium iodide (red)

Marker gene tethering by nucleoporins affects gene expression in plants

In non-plant systems, chromatin association with the nuclear periphery affects gene expression, where interactions with nuclear envelope proteins can repress and interactions with nucleoporins can enhance transcription. In plants, both hetero- and euchromatin can localise at the nuclear periphery, but the effect of proximity to the nuclear periphery on gene expression remains largely unknown. This study explores the putative function of Seh1 and Nup50a nucleoporins on gene expression by using the Lac Operator / Lac Repressor (LacI-LacO) system adapted to Arabidopsis thaliana. We used LacO fused to the luciferase reporter gene (LacO:Luc) to investigate whether binding of the LacO:Luc transgene to nucleoporin:LacI protein fusions alters luciferase expression. Two separate nucleoporin-LacI-YFP fusions were introduced into single insert, homozygous LacO:Luc Arabidopsis plants. Homozygous plants carrying LacO:Luc and a single insert of either Seh1-LacI-YFP or Nup50a-LacI-YFP were tested for luciferase activity and compared to plants containing LacO:Luc only. Seh1-LacI-YFP increased, while Nup50a-LacI-YFP decreased luciferase activity. Seh1-LacI-YFP accumulated at the nuclear periphery as expected, while Nup50a-LacI-YFP was nucleoplasmic and was not selected for further study. Protein and RNA levels of luciferase were quantified by western blotting and RT-qPCR, respectively. Increased luciferase activity in LacO:Luc+Seh1-LacI-YFP plants was correlated with increased luciferase protein and RNA levels. This change of luciferase expression was abolished by disruption of LacI-LacO binding by treating with IPTG in young seedlings, rosette leaves and inflorescences. This study suggests that association with the nuclear periphery is involved in the regulation of gene expression in plants

A novel family of plant nuclear envelope associated proteins

This paper describes the characterisation of a new family of higher plant nuclear envelope associated proteins (NEAPs) that interact with proteins of the nuclear envelope. In the model plant Arabidopsis thaliana, the family consists of three genes expressed ubiquitously (AtNEAP1-3) and a pseudogene (AtNEAP4). NEAPs consist of extensive coiled-coil domains, followed by a nuclear localisation signal and a C-terminal predicted transmembrane domain. Domain deletion mutants confirm the presence of a functional nuclear localisation signal and transmembrane domain. AtNEAP proteins localise to the nuclear periphery as part of stable protein complexes, are able to form homo- and heteromers and interact with the SUN domain proteins AtSUN1 and AtSUN2, involved in the Linker of Nucleoskeleton and Cytoskeleton (LINC) complex. An A. thaliana cDNA library screen identified a putative transcription factor called AtbZIP18 as a novel interactor of AtNEAP1, which suggest a connection between NEAP and chromatin. An Atneap1 Atneap3 double knock out mutant showed reduced root growth and altered nuclear morphology and chromatin structure. Thus AtNEAPs are suggested as INM anchored coiled-coil proteins with roles in maintaining nuclear morphology and chromatin structure